The long term objective of this grant application is to identify, map, sequence and define in molecular terms the biochemical and biophysical changes that mediate the action of the human hormone granulocyte-- macrophage colony-stimulating factor on human neutrophils. This growth factor, which is released by several activated cells such as T lymphocytes, is an important stimulus for the proliferation of erythroid and myelomonocytic stem cells in vitro . In patients with AIDS, it increases the number of neutrophils, eosinophils and monocytes. It is also an important in vivo regulator of granulopoiesis and neutrophil function; it induces significant leukocytosis and shortens the period of neutropenia following bone marrow transplantation. The addition of this growth factor to mature human neutrophils primes these cells to subsequent stimulation by the chemotactic factor fMet-Leu-Phe. Thus, it plays an important role in the host defense. In spite of its importance very little is known about the mechanism of its action. In this work, particular emphasis will be placed upon delineating the differences in the action of GM-CSF and other stimuli on neutrophils in suspension versus attached to surface. Several techniques will be used and they include, among others, protein phosphorylation, superoxide generation, immunoblotting, binding, ADP-ribosylation, generation of phosphatidic acid and production of diacylglycerol.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI028810-03
Application #
2064647
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1992-04-01
Project End
1996-03-31
Budget Start
1994-04-01
Budget End
1996-03-31
Support Year
3
Fiscal Year
1994
Total Cost
Indirect Cost
Name
University of Connecticut
Department
Anatomy/Cell Biology
Type
Schools of Dentistry
DUNS #
City
Farmington
State
CT
Country
United States
Zip Code
06030
Waterman, W H; Molski, T F; Huang, C K et al. (1996) Tumour necrosis factor-alpha-induced phosphorylation and activation of cytosolic phospholipase A2 are abrogated by an inhibitor of the p38 mitogen-activated protein kinase cascade in human neutrophils. Biochem J 319 ( Pt 1):17-20
Nahas, N; Molski, T F; Fernandez, G A et al. (1996) Tyrosine phosphorylation and activation of a new mitogen-activated protein (MAP)-kinase cascade in human neutrophils stimulated with various agonists. Biochem J 318 ( Pt 1):247-53
Nahas, N; Waterman, W H; Sha'afi, R I (1996) Granulocyte-macrophage colony-stimulating factor (GM-CSF) promotes phosphorylation and an increase in the activity of cytosolic phospholipase A2 in human neutrophils. Biochem J 313 ( Pt 2):503-8
Gomez-Cambronero, J (1995) Immunoprecipitation of a phospholipase D activity with antiphosphotyrosine antibodies. J Interferon Cytokine Res 15:877-85
Waterman, W H; Sha'afi, R I (1995) A mitogen-activated protein kinase independent pathway involved in the phosphorylation and activation of cytosolic phospholipase A2 in human neutrophils stimulated with tumor necrosis factor-alpha. Biochem Biophys Res Commun 209:271-8
Waterman, W H; Sha'afi, R I (1995) Effects of granulocyte-macrophage colony-stimulating factor and tumour necrosis factor-alpha on tyrosine phosphorylation and activation of mitogen-activated protein kinases in human neutrophils. Biochem J 307 ( Pt 1):39-45
Fouda, S I; Molski, T F; Ashour, M S et al. (1995) Effect of lipopolysaccharide on mitogen-activated protein kinases and cytosolic phospholipase A2. Biochem J 308 ( Pt 3):815-22
Johnson, G M; Gomez-Cambronero, J (1995) Priming of tyrosine phosphorylation in GM-CSF-stimulated adherent neutrophils. J Leukoc Biol 57:692-8
Durstin, M; Gao, J L; Tiffany, H L et al. (1994) Differential expression of members of the N-formylpeptide receptor gene cluster in human phagocytes. Biochem Biophys Res Commun 201:174-9
Durstin, M; Durstin, S; Molski, T F et al. (1994) Cytoplasmic phospholipase A2 translocates to membrane fraction in human neutrophils activated by stimuli that phosphorylate mitogen-activated protein kinase. Proc Natl Acad Sci U S A 91:3142-6

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